Hormone secretion by exocytosis with emphasis on information from the chromaffin cell system.
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Animals↗
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Presence of cellular renin-angiotensin system in chromaffin cells of bovine adrenal medulla.
The presence of a local renin-angiotensin system has been established in organs that serve as angiotensin targets. In this study, the expression of angiotensinogen mRNA and subcellular localization of renin, angiotensin-converting enzyme, and angiotensin II were investigated in bovine adrenal medullary cells in primary culture. By light microscopy, expression of angiotensinogen mRNA, immunoreactive renin, angiotensin-converting enzyme, and angiotensin II were readily detectable only in the chromaffin cells. The density distribution of renin and angiotensin II in sucrose gradients suggested a concentration in chromaffin granules, a localization directly confirmed by immunoelectron microscopy. Reverse transcriptase-polymerase chain reaction and sequencing confirmed the expression of angiotensinogen in bovine chromaffin cells and the adrenal medulla. In addition, in vitro autoradiography indicated that both angiotensin-converting enzyme and angiotensin type 1 receptors were present in the adrenal medulla. These results provide the first direct evidence that chromaffin cells in the adrenal medulla are not only the target for angiotensin but should also be considered as potential local angiotensin-generating and -storing cells.
Adrenal Medulla↗
On the appearance of monoamines in the sympathetic systems and the chromaffin tissue in the mouse embryo.
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Amines↗
Deletion of tyrosine hydroxylase gene reveals functional interdependence of adrenocortical and chromaffin cell system in vivo.
Catecholamines are produced in the medulla of the adrenal gland and may participate in the intraglandular regulation of its cortex. We analyzed the adrenal structure and function of albino tyrosine hydroxylase-null (TH-null) mice that are deficient in adrenal catecholamine production. Adrenal catecholamines were markedly reduced, and catecholamine histofluorescence was abrogated in 15-day-old TH-null mice. Chromaffin cell structure was strikingly altered at the ultrastructural level with a depletion of chromaffin vesicles and an increase in rough endoplasmic reticulum compared with wild-type mice. Remaining chromaffin vesicles lined up proximally to the cell membrane in preparation for exocytosis providing a "string-of-pearls" appearance. There was a 5-fold increase in the expression of proenkephalin mRNA (502.8 +/- 142% vs. 100 +/- 17.5%, P = 0.016) and a 2-fold increase in the expression of neuropeptide Y (213.4 +/- 41.2% vs. 100 +/- 59.9%, P = 0.014) in the TH-null animals as determined by quantitative TaqMan (Perkin-Elmer) PCR. Accordingly, immunofluorescence for met-enkephalin and neuropeptide tyrosine in these animals was strongly enhanced. The expression of phenylethanolamine N-methyl transferase and chromogranin B mRNA was similar in TH-null and wild-type mice. In TH-null mice, adrenocortical cells were characterized by an increase in liposomes and by tubular mitochondria with reduced internal membranes, suggesting a hypofunctional state of these steroid-producing cells. In accordance with these findings, plasma corticosterone levels were decreased. Plasma ACTH levels were not significantly different in TH-null mice. In conclusion, both the adrenomedullary and adrenocortical systems demonstrate structural and functional changes in catecholamine-deficient TH-null mice, underscoring the great importance of the functional interdependence of these systems in vivo.
Adrenal Glands↗
Glomus faciale, glomus jugulare, glomus tympanicum, glomus vagale, carotid body tumors, and simulating lesions. Role of MR imaging.
In summary, MR imaging characteristics of a case of paraganglioma of the facial nerve are reported. The relationship of paragangliomas and the chromaffin system have been discussed. There are many reports of cases of synchronous paragangliomas and pheochromocytomas. These reports, along with simultaneous involvement in familial MEN syndromes, and the common embrylogic origin (neural crest) and similar histopathologic relationships between paragangliomas and pheochromocytoma, all support the fact that they are part of the chromaffin system.
Adrenal Gland Neoplasms↗
Trophic factors from chromaffin granules promote survival of peripheral and central nervous system neurons.
Chromaffin cells of the adrenal medulla were used to study the release of neurotrophic factors operationally defined by their capacity to promote the in vitro survival of embryonic neurons from the peripheral and central nervous system. Chromaffin cells are closely related to sympathetic neurons in terms of their transmitters and specific proteins and, like sympathetic neurons, receive preganglionic cholinergic, aminergic and peptidergic neuronal inputs. The issue of whether chromaffin cells store and secrete neurotrophic factors is therefore pertinent to the question whether trophic mechanisms may be involved in neuronal interactions and what modes of secretion are employed to liberate neurotrophic factors from neurons. Cell culture media conditioned by purified bovine chromaffin cells supported several neuron populations in vitro. Stimulation of the chromaffin cells with the cholinergic agonist carbachol (10(-4) M) increased in parallel the output of neurotrophic factor activity (assayed on chick ciliary ganglionic neurons) as well as two components specifically located in chromaffin granules, chromogranin A and catecholamines. The release of all three components was partially blocked by the Ca2+ channel blocker verapamil (10(-5) M), suggesting co-storage and -release of neurotrophic factors, chromogranin A and catecholamines in/from chromaffin granules. Neurotrophic factor activity for ciliary ganglionic neurons accumulating in the medium of unstimulated chromaffin cells decreased with time, and so did catecholamines. In contrast, amounts of neurotrophic factors and catecholamines released by challenging cells with carbachol did not significantly decline up to 62 h. The neurotrophic factor activity tested on chick ciliary, sensory and spinal cord neurons as well as on rat hippocampal neurons was heat- and trypsin-labile and could not be blocked by polyclonal antibodies against bovine nerve growth factor and the chromogranin A, B, and C. Defined fragments of chromogranin A and pancreastatin were devoid of neurotrophic activity. Our results suggest the presence of one or several neurotrophic factors in chromaffin granules, which can be released by exocytosis and may be potentially relevant for the maintenance of neurons innervating the adrenal medulla.
Adrenal Medulla↗
URINARY EXCRETION OF CATECHOL AMINES IN THE RAT AFTER THEIR LIBERATION BY RESERPINE OR DEXAMPHETAMINE.
Daily urinary excretion of catechol amines in normal rats and in rats from which the adrenal medullae had been removed has been determined by a photofluorimetric method. In both groups, reserpine (2 mg/kg, intraperitoneally) produces: (1) A decrease in the urinary excretion of noradrenaline which persists for more than 3 weeks; this action is not influenced by monoamine oxidase inhibitors and mecamylamine. (2) An increase, within 20 to 68 hr, in the urinary excretion of adrenaline, even though the urine of rats without adrenal medullae does not usually contain adrenaline. These effects are prevented by monoamine oxidase inhibitors and, in the normal animals, are reduced by mecamylamine. In both groups, dexamphetamine (6 mg/kg, intraperitoneally) produces an increase in the excretion of adrenaline and noradrenaline, the adrenaline appearing in the urine of the rats without adrenal medullae within 20 to 44 hr. Mecamylamine prevents the effect of dexamphetamine on the excretion of noradrenaline. Dexamphetamine, administered within a week of reserpine treatment, produces its usual effects on the urinary excretion of catechol amines in normal rats, but has no effect in rats without adrenal medullae. The results are discussed with regard to both the mechanism by which reserpine and dexamphetamine influence the peripheral stores of adrenaline and noradrenaline, and the significance of the adrenal and extra-adrenal chromaffin system.
Adrenal Medulla↗
Catecholamine release from bovine adrenal chromaffin cells during anoxia or metabolic inhibition.
A significant release of catecholamines within the heart has been observed during myocardial ischemia. Because this can be markedly inhibited by amine-uptake-blocking agents, it has been suggested that its mechanism is a carrier-mediated efflux from neurons, which is not operative under normal conditions. The present work examined this release process in chromaffin cells isolated from the bovine adrenal medulla, a model system for studying the sympathetic nervous system. Chromaffin cells in primary culture retained normal secretory responses for up to 7 days. Conditions designed to mimic ischemia, that is, anoxia or metabolic inhibition, resulted in a significant release of catecholamines. This release was shown to be independent of extracellular calcium but, in contrast to the release observed in ischemic hearts, was not inhibited by amine-uptake blockers. Electrophoresis with immunoblotting demonstrated that significant levels of the chromaffin granule protein, chromogranin A, were released during metabolic inhibition, indicative of an exocytotic mechanism. However, there was no release of the cytosolic protein, lactate dehydrogenase, indicating that there was no concomitant breakdown of the cell membrane. These results provide evidence for an exocytotic release of catecholamines mediated by the direct action of conditions of metabolic inhibition.
Adrenal Medulla↗
Solubilization and reconstitution of the adenosine 5'-triphosphate dependent noradrenaline uptake system of bovine chromaffin granule membrane.
The ATP-dependent catecholamine uptake system of chromaffin granule membrane has been solubilized and reconstituted in phospholipidic vesicles. The activity of the vesicles implies that both the ATP-dependent H+-translocase and the noradrenaline carrier have been successfully reconstituted. The membrane was solubilized by sodium cholate in presence of asolectin and the asolectin to cholate ratio appeared to be critical. Omission of asolectin resulted in reconstitution of vesicles with an active H+-pump and an inactive transport system. The detergent was removed from the solubilized membranes by filtration on Sephadex G-50 and it has been verified that the residual detergent of the reconstituted preparation was below the concentration inhibitory to the ghost H+-pump. The pH-dependence, Km for ATP and Km for noradrenaline of the reconstituted vesicles were similar to those of the ghosts, but their specific activity and reserpine-resistance were somehow variable. Vesicle activity was limited by transporter reconstitution, thus suggesting that reconstitution of the complete system might be used as an assay for the transporter. The noradrenaline carrier is not physically linked to dopamine beta-hydroxylase and bears no wheat germ agglutinin binding sites.
Adenosine Triphosphate↗
Glucocorticoid receptors and regulation of phenylethanolamine-N-methyltransferase activity in cultured chromaffin cells.
Glucocorticoids are known to regulate the enzyme phenylethanolamine-N-methyltransferase (PNMT) in the adrenal medulla of the rat and are thereby thought to control the synthesis of epinephrine. We have examined the details of this relationship in a simplified system, chromaffin cell primary cultures derived from bovine adrenal medulla. Cultured chromaffin cells were found to have a cytosolic, high affinity, saturable glucocorticoid-binding protein with the steroid specificity of a classical glucocorticoid receptor and a Kd of approximately 1 nM. Treatment of cultured cells with dexamethasone or hydrocortisone at any time up to 21 days in culture increased PNMT activity in the soluble fraction of the cell. The concentration of hormone required to produce a half-maximal response was 10 nM dexamethasone when cells were cultured in the presence of 5% fetal calf serum, or 1 nM in a defined serum-free medium. These dose-response relationships are consistent with mediation of this effect by the glucocorticoid receptor. Unexpectedly, however, the glucocorticoid-induced increment in PNMT activity was not inhibited by cycloheximide at concentrations up to 50 microM, and an acceleration of protein synthesis by insulin treatment did not augment the glucocorticoid effect on PNMT. Treatment of the cells with dexamethasone (100 microM) prevented the decline in the epinephrine-to-norepinephrine ratio seen over time in culture, an effect consistent with increased PNMT activity. However, there was no effect of dexamethasone on the ability of the cells to secrete catecholamines in response to stimulation with high KCl or 30 microM nicotine.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Identification of chromaffin granule-binding proteins. Relationship of the chromobindins to calelectrin, synhibin, and the tyrosine kinase substrates p35 and p36.
At least 23 soluble proteins (chromobindins) bind to chromaffin granule membranes in the presence of Ca2+. In order to further the identification of the chromobindins and to determine the roles they may play in exocytosis or other aspects of chromaffin cell biology, several of these proteins were compared to other known membrane-binding proteins. Chromobindin 4 was identified as a 32-kDa protein called calelectrin or endonexin. Immunologically related proteins were detected in bovine brain and human platelets. Chromobindin 20 was identified as a 67-kDa variant of calelectrin and was found to have the activities of the synexin inhibitory protein, synhibin. Chromobindin 8 was identified as p36, a substrate for the tyrosine-specific kinase, pp60v-src. Chromobindin 8 was also demonstrated to undergo phosphorylation predominantly on alkali-sensitive sites during stimulation of the chromaffin cell with 20 microM nicotine. Chromobindin 6 was identified as p35, a substrate for the tyrosine kinase activity associated with the epidermal growth factor receptor. Chromobindin 9, which is known to be a substrate for protein kinase C (Ca2+/phospholipid-dependent enzyme), was found to be immunologically related to p35 and may be a precursor of chromobindin 6. The identification of these proteins from the chromaffin system may be useful in the characterization of similar, complex groups of membrane-binding proteins that have been observed in other systems.
Amino Acids↗
Exocytosis studies in a chromaffin cell-free system: imaging of single-vesicle exocytosis in a chromaffin cell-free system using total internal reflection fluorescence microscopy.
We have developed a system for the real-time study of regulated exocytosis in living, cultured bovine adrenal chromaffin cells (BCCs). Exocytosis was monitored by the use of total internal reflection fluorescence (TIRF) microscopy to image single large dense-core secretory vesicles (LDCVs). Fluorescent labeling of LDCVs was achieved either with the membrane-permeant weak base, acridine orange (AO), or by transduction of BCCs so as to express a fluorescent chimeric "cargo" protein that is targeted to LDCVs. In either case, exocytosis is visible by the disappearance of a vesicle accompanied by a bright flash as the fluorescent contents leave the acidic LDCV lumen, move towards the source of the evanescent wave, and disperse into the extracellular medium. Furthermore, for the first time, we have developed a broken-cell system for real-time imaging in BCCs, in which individual plated cells are mechanically "unroofed" with a jet of intracellular medium, leaving a membrane patch with docked vesicles on the coverslip. In this cell-free system, a subpopulation of docked granules undergoes exocytosis in response to calcium. This approach allows us direct experimental access to membrane-docked LDCVs in order to investigate the dependence of exocytosis on defined protein components and intracellular conditions at the single-vesicle level. In addition, this system can be used for a reconstitution analysis of the exocytosis machinery. Finally, we demonstrate the use of 2D+1 image analysis for visualizing single-vesicle exocytosis. We use this approach for a rapid analysis of larger numbers of imaged vesicles.
Acridine Orange↗
Chromaffin tissue of the sympathetic ganglia and heart.
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Chromaffin System↗
CATECHOLAMINE-CONTAINING STRUCTURES IN THE HYPOGASTRIC NERVES OF THE DOG.
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Catecholamines↗
The local chromaffin cell plasminogen/plasmin system and the regulation of catecholamine secretion.
Chromaffin cells express components of the plasminogen/plasmin system, including its major activator, tissue plasminogen activator (t-PA), and high-affinity cellular receptors for plasminogen, which promote local concentration and activation of plasminogen at the cell surface. Our studies suggest that plasmin participates in local neuroendocrine prohormone processing and that perturbation of this system profoundly affects the secretory characteristics of the cells. These results suggest the presence of a local, functionally active, chromaffin cell plasminogen/plasmin system that plays a major role in the regulation of catecholamine release from catecholaminergic cells.
Animals↗
A new focus on interoceptive properties of adrenal medulla.
The adrenal medulla is an important part of the sympathoadrenal system. Chromaffin cells of the adrenal medulla respond to a broad spectrum of stressful situations by releasing epinephrine and norepinephrine. Originally, it was accepted that this response is controlled exclusively by central nervous system structures. However, it was also demonstrated that a surgically denervated adrenal medulla can respond directly by secreting epinephrine and norepinephrine during an imbalance of internal environment (hypoglycemia, asphyxia). Published data had documented the innervation of the adrenal medulla by sensory neurons of spinal dorsal root ganglia. In addition, recent data showed that ganglion cells of the adrenal medulla project ascending axons. These data suggested potential transmission of information from the adrenal medulla to the central nervous system regarding metabolic changes in the blood. This paper presents an overview of possible involvement of adrenal medullary chromaffin cells in the detection of changes in the internal environment and in the transmission of this information to the central nervous system.
Adrenal Medulla↗
Retroperitoneal paraganglia and the peripheral autonomic nervous system in the human fetus.
By means of the AChE in toto staining method retroperitoneal paraganglia and the peripheral autonomic nervous system in human fetuses have been investigated. Many small retroperitoneal paraganglia are present near the sympathetic trunks close to the sympathetic trunk ganglia. In the thoracic region small paraganglia are present in the intercostal spaces. Small splanchnic nerves entering small paraganglia have been described. In the lower sacral region no paraganglia are present. The major splanchnic nerve arises at various levels from the sympathetic trunks as well as many smaller thoracic splanchnic nerves. Intermediate ganglia are present in the major splanchnic nerve, the smaller splanchnic nerves and the communicating rami. In the sympathetic trunks many ganglia are fused. In the human fetus there exists a large variability in number and diameter of the communicating rami. Interconnecting bundles of nerve fibers between the left and right sympathetic trunks are present at all levels, but most numerous at the sacral level.
Autonomic Nervous System↗
[Adrenal medullitis in cases of human rabies].
The adrenal glands and the central nervous system were studied from five human cases of rabies who died three to six days after first rings of clinical manifestations were seen. In all cases there were Negri bodies in the cytoplasm of neurons of the central nervous system and mononuclear inflammatory cells around blood vessels, more intense in the cases with longer survival. Only the adrenal medulla showed diffuse and intense mononuclear exudate associated with pheocromocytes alterations in 60% of patients. Eosinophilic bodies were found in the cytoplasm of pheochromocytes and in the interstitial space. These bodies may be similar to Negri bodies but their true nature it is not known. The pathogenesis of adrenal medullitis in rabies may be related to embryological and metabolical relationships of the chromaffin system and the nervous system.
Adrenal Gland Diseases↗